A series of novel oxazolo[5,4-d]pyrimidines was designed via a scaffold hopping strategy and synthesized through a newly developed approach. All these compounds were evaluated for their biological activity toward CB₁/CB₂ cannabinoid receptors, their metabolic stability in mice liver microsomes and their cytotoxicity against several cell lines. Eight compounds have been identified as CB₂ ligands with K_i values less than 1 μM. It is noteworthy that 2-(2-chlorophenyl)-5-methyl-7-(4-methylpiperazin-1-yl) oxazolo[5,4-d]pyrimidine 47 and 2-(2-chlorophenyl)-7-(4-ethylpiperazin-1-yl)- 5-methyloxazolo[5,4-d]pyrimidine 48 showed CB₂ binding affinity in the nanomolar range and significant selectivity over CB₁ receptors. Interestingly, functionality studies imply that they behave as competitive neutral antagonists. Moreover, all tested compounds are devoid of cytotoxicity toward several cell lines, including Chinese hamster ovary cells (CHO) and human colorectal adenocarcinoma cells HT29.